The purpose of these studies is to generate and characterize monoclonal antibodies that are reactive with human mammary tumor cells. Splenic lymphocytes of mice, immunized with membrane-enriched fractions of mestastatic human mammary carcinoma tissues, were fused with murine myeloma cells. This resulted in the generation of hybridoma cultures secreting Igs reactive in solid phase radioimmunoassays with extracts of metastatic mammary carcinoma cells from involved livers, but not to extracts of apparently normal human tissues. Eleven monoclonal antibodies were chosen which demonstrated reactivities with human mammary tumor cells and not with apparently normal human mammary tissue. These monoclonals could be placed into at least five major groups based on their differential binding to the surface of various live human mammary tumor cells in culture, to extracts of mammary tumor tissues, or to tissue sections of mammary tumor cells employing the immunoperoxidase technique. Monoclonal antibodies of all five major groups demonstrated binding to human metastatic mammary carcinoma cells both in axillary lymph nodes and at distal sites.